Analyte sensor and analyte sensing method

ABSTRACT

A biosensor includes a detection element having an analyte detecting portion which is monotonically increased in mass in response to detection of an analyte; a reference element having a reference measuring portion which exhibits no reactivity to the analyte; a mixer which mixes a detection signal responsive to mass variations in the analyte detecting portion from the detection element and a reference signal from the reference element; a measurement portion which calculates two candidate phase-change values of a positive value and a negative value, from a signal mixed by the mixer in accordance with a heterodyne system, and determines a phase-change value from the two candidate phase change value by judging whether the phase is positive or negative based on temporal changes in signal strength; and a detection amount calculation portion which calculates a detection amount of the analyte based on the phase change value determined by the measurement portion.

CROSS REFERENCE TO RELATED APPLICATIONS

The present application is a divisional of U.S. application Ser. No.14/375,780 filed on Jul. 30, 2014, which is the national stage of PCTApplication No. PCT/JP2013/051887 filed on Jan. 29, 2013, which claimsthe benefit of Japanese Application Nos. 2012-016383, filed on Jan. 30,2012, and 2012-074156, filed on Mar. 28, 2012. The contents of each ofthe above applications is incorporated herein by reference in itsentirety.

TECHNICAL FIELD

The present invention relates to an analyte sensor capable of measuringproperties of an analyte or a target substance contained in an analyteas well as to an analyte sensing method.

BACKGROUND ART

There is known a surface acoustic wave sensor for measuring propertiesor ingredients of an analyte liquid by means of a surface acoustic wavedevice.

The surface acoustic wave sensor, which is constructed of apiezoelectric substrate on which is mounted a detecting portion whichreacts with a component contained in an analyte sample, is designed todetect the properties or ingredients of an analyte liquid by measuringelectric signals responsive to variations in surface acoustic wave (SAW)propagating through the detecting portion (for example, refer to PatentLiterature 1).

The SAW sensor disclosed in Patent Literature 1 measures theconcentration of an analyte sample by detecting a phase difference inSAW. In order to make phase difference measurement, a quadraturemodulation system has generally been adopted, because it offers anextended measurable phase range.

CITATION LIST Patent Literature

Patent Literature 1: Japanese Unexamined Patent Publication JP-A2008-122105

Non-Patent Literature

Non-Patent Literature 1: “Development of Novel SAW Liquid Sensing Systemwith SAW Signal Generator” excerpted from the technical report of IEICEpublished in February, 2003 by The Institute of Electronics, Informationand Communication Engineers

SUMMARY OF INVENTION Technical Problem

However, the quadrature modulation system poses the following problems:the number of components constituting the system is large with aconsequent difficulty in system downsizing; and the number of digitalprocessing steps is large with a consequent increase in currentconsumption.

In light of this, there has been a demand for small-scale dataprocessing method featuring lower current consumption and a biosensorequipped with the data processing method.

Solution to Problem

According to one aspect of the invention, an analyte sensor comprises:an analyte detecting portion; a detection element; a reference measuringportion; a reference element; a measurement portion; and a detectionamount calculation portion. The analyte detecting portion ismonotonically changed in mass in response to adsorption of a targetprovided in an analyte or reaction with the target. The detectionelement is configured to output a detection signal of AC responsive tomass variations in the analyte detecting portion. The referencemeasuring portion undergoes neither adsorption of a target nor reactionwith the target. The reference element is configured to output areference signal of AC relative to the detection signal. The measurementportion determines two candidate phase-change values of a positivecandidate phase-change value and a negative candidate phase-changevalue, from a measurement signal which is obtained from the detectionsignal and the reference signal in accordance with a heterodyne system.At this time, case classification is carried out according to thefollowing four conditions (1) to (4):

(1) where a mass of the analyte detecting portion is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system;

(2) where the mass of the analyte detecting portion is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system;

(3) where the mass of the analyte detecting portion is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system; and

(4) where the mass of the analyte detecting portion is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system.

Under the condition (1) or (2), when measurement signal strength isdecreased with time, the positive candidate phase-change value isoutputted as a phase change value, and when measurement signal strengthis increased with time, the negative candidate phase-change value isoutputted as a phase change value.

Under the condition (3) or (4), when measurement signal strength isdecreased with time, the negative candidate phase-change value isoutputted as a phase change value, and when measurement signal strengthis increased with time, the positive candidate phase-change value isoutputted as a phase change value.

The detection amount calculation portion calculates the detection amountof the analyte on the basis of the phase change value.

According to one aspect of the invention, an analyte sensing methodcomprises: an analyte solution supplying step; a determination step; anda calculation step. In the analyte solution supplying step, an analytesolution containing an analyte in which a target is provided, issupplied to an analyte detecting portion of a detection element that ismonotonically changed in mass in response to adsorption of the target orreaction with the target, and a reference detecting portion of areference element that undergoes neither adsorption of the target norreaction with the target. In the determination step, two candidatephase-change values of a positive candidate phase-change value and anegative candidate phase-change value, are determined from a measurementsignal which is obtained from a detection signal of AC responsive tomass variations in the analyte detecting portion and a reference signalof AC relative to the detection signal from the reference detectingportion, in accordance with a heterodyne system. At this time, caseclassification is carried out according to the following four conditions(1) to (4):

(1) where a mass of the analyte detecting portion is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system;

(2) where the mass of the analyte detecting portion is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system;

(3) where the mass of the analyte detecting portion is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system; and

(4) where the mass of the analyte detecting portion is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system.

Under the condition (1) or (2), when measurement signal strength isdecreased with time, the positive candidate phase-change value isdetermined as a phase change value, and when measurement signal strengthis increased with time, the negative candidate phase-change value isdetermined as a phase change value.

Likewise, under the condition (3) or (4), when measurement signalstrength is decreased with time, the negative candidate phase-changevalue is determined as a phase change value, and, when the measurementsignal strength is increased with time, the positive candidatephase-change value is determined as a phase change value.

In the calculation step, the amount of the analyte detected iscalculated on the basis of the phase change value.

Advantageous Effects of Invention

According to the invention, it is possible to provide a compact analytesensor featuring lower current consumption and an analyte sensingmethod.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a principled configuration diagram showing an analyte sensorin accordance with an embodiment of the invention;

FIG. 2 is a perspective view of the analyte sensor in accordance withthe embodiment of the invention;

FIG. 3 is a partially cutaway perspective view of the analyte sensorshown in FIG. 1;

FIG. 4A is a sectional view taken along the line IVa-IVa shown in FIG.2, and FIG. 4B is a sectional view taken along the line IVb-IVb shown inFIG. 2;

FIG. 5 is a top view of the analyte sensor shown in FIG. 1, with partthereof removed;

FIGS. 6A and 6B show a modified example of the analyte sensor shown inFIG. 1;

FIGS. 7A and 7B show a modified example of the analyte sensor shown inFIG. 1;

FIG. 8 shows a modified example of the analyte sensor shown in FIG. 1;

FIG. 9 is a principled configuration diagram showing the analyte sensorin accordance with another embodiment of the invention;

FIG. 10 is a graph showing a correlation between detection signalstrength and phase in the analyte sensor shown in FIG. 1;

FIG. 11 is a chart showing a correlation between measurement time andphase difference in the example of the invention; and

FIG. 12 is a chart for comparison between the example of the inventionand VNA.

DESCRIPTION OF EMBODIMENTS

Hereinafter, embodiments of an analyte sensor pursuant to the inventionwill be described in detail with reference to drawings. Note that, ineach of the drawings as will hereafter be described, identicalconstituent members are identified with the same reference symbols.Moreover, the size of each member, the distance between the members, andso forth are schematically depicted and may therefore be different fromthe actual measurements.

Moreover, although any side of the analyte sensor may be either an upperside or a lower side, in the following description, for purposes ofconvenience, an x-y-z rectangular coordinate system is defined, and,words such as an upper surface, a lower surface, etc. are used on theunderstanding that a positive z direction is an upward direction.

Principles of Analyte Sensor 100

FIG. 1 is a schematic diagram for explaining the principle of an analytesensor 100. As shown in FIG. 1, the analyte sensor 100 comprises adetection element 110; a reference element 120; a measurement portion140; and a detection amount calculation portion 150.

In this embodiment, the detection element 110 includes an analytedetecting portion 111 which is monotonically increased in mass inresponse to adsorption of a target provided in an analyte or reactionwith the target. For example, the analyte detecting portion 111 can beimplemented by immobilizing a reactive group having such a reactivity asto undergo specific target adsorption on a Au film impervious to theinfluence of electrical characteristics such as electrical conductivityof an analyte. Note that there is no need for the analyte detectingportion to adsorb a target in itself. For example, a reactive grouphaving such a characteristic as to react with a substance other than atarget provided in an analyte resulting from reaction with the targetmay be immobilized on a Au film. It is preferable that this Au film iselectrically short-circuited.

The reference element 120 includes a reference measuring portion 121.For example, the reference measuring portion 121 does not have such areactivity as to specifically adsorb a target provided in an analyte orto cause substitution reaction with a substance contained in an analytedue to a conformational change. As a concrete example, a Au film freefrom immobilization of the aforementioned reactive group can be used.

A mixer 130 mixes a detection signal responsive to mass variations inthe analyte detecting portion 111 from the detection element 110 and areference signal from the reference element 120. In this case, thedetection signal and the reference signal are AC signals, and, thereference signal serves as a fiducial signal in relation to thedetection signal.

The measurement portion 140 performs two steps as set forth hereunder.At first, from a measurement signal mixed by the mixer 130, through alow-pass filter 131, a candidate phase-change value is calculated inaccordance with a heterodyne system. The processing details of this stepvary depending on the following four conditions. Concretely, caseclassification is carried out according to the following four conditions(1) to (4):

(1) where the mass of the analyte detecting portion 111 is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system;

(2) where the mass of the analyte detecting portion ill is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system;

(3) where the mass of the analyte detecting portion 111 is monotonicallyincreased, and the detection signal is obtained by subtracting thereference signal from the detection signal in accordance with theheterodyne system; and

(4) where the mass of the analyte detecting portion 111 is monotonicallydecreased, and the detection signal is obtained by subtracting thedetection signal from the reference signal in accordance with theheterodyne system.

In the case where the mass of the analyte detecting portion 111 ismonotonically increased, a detection signal falls behind in phase ascompared to a reference signal. Therefore, when a detection signal issubtracted from a reference signal, there is a monotonic increase inphase change. On the other hand, when a reference signal is subtractedfrom a detection signal, there is a monotonic decrease in phase change.

Likewise, in the case where the mass of the analyte detecting portion111 is monotonically decreased, a detection signal advances in phase ascompared to a reference signal. Therefore, when a detection signal issubtracted from a reference signal, there is a monotonic decrease inphase change. On the other hand, when a reference signal is subtractedfrom a detection signal, there is a monotonic increase in phase change.This phenomenon is utilized.

In giving a more concrete explanation, the condition (1) will be quotedby way of illustration. Since a measurement signal is processed by theheterodyne system, as shown in FIG. 10, the signal makes a sine curvewhich is positive-negative symmetrical about 0°, wherefore a candidatephase-change value corresponding to signal strength (output value) y1takes on two negative and positive values, namely x1 and x2. Herein,FIG. 10 shows a locus curve indicative of a correlation betweenmeasurement signal strength and phase. Next, temporal changes in thesignal mixed by the mixer 130 are confirmed. Since the analyte detectingportion 111 has the characteristic of being monotonically increased inmass, it follows that the phase of the measurement signal ismonotonically increased with time. With the utilization of thischaracteristic, it will be found that, when the measurement signalstrength is increased, the negative phase value x1 is assigned, and whenthe measurement signal strength is decreased, the positive phase valuex2 is assigned. In other words, while signal strength varies with time(Δx), when the phase takes on the value x1, the strength of x1+Δx islarger than y1. On the other hand, when the phase takes on the value x2,the strength of x2+Δx is smaller than y1. Thus, a distinction can bemade between x1 and x2 by determining whether the phase value becomeslarger or smaller than the original strength (y1) with time. That is,whether the phase is positive or negative can be judged by examiningtemporal changes in the measurement signal mixed by the mixer 130. Inthis way, a decision is made between the two candidate phase-changevalues to determine a phase change value. Also in the case of thecondition (2), a phase change value can be determined in a similarmanner. By contrast, in the cases of the conditions (3) and (4), whenthe measurement signal strength is increased, the positive phase valueis determined as the phase change value, and when the measurement signalstrength is decreased, the negative phase value is determined as thephase change value.

Then, the detection amount calculation portion 150 calculates thedetection amount of an analyte on the basis of the phase change valuedetermined by the measurement portion 140.

By virtue of such a configuration, an analyte sensor 100 capable ofcalculating the detection amount of an analyte can be provided. In thisconstruction, since signal processing operation is performed by theheterodyne system, it is possible to calculate the detection amount ofan analyte only with the addition of the mixer 130 for deriving adifferential between a detection signal and a reference signal.Accordingly, in contrast to the case of adopting the quadraturemodulation system as has been conventionally used, the analyte sensor100 does not necessitate complicated signal processing operation, hasfewer necessary components, can be made lower in profile, and succeedsin reduction of current consumption. Moreover, in a normal heterodynesystem, a judgment as to whether the phase is positive or negativecannot be made, wherefore measurable phases are limited to a range from0° to 180°. However, according to the analyte sensor 100 of the presentembodiment, by confirming temporal changes in signal, whether the phaseis positive or negative can be judged from candidate phase-changevalues. This makes it possible to obtain a wider measurable phase rangeextending from −180° to 180°. Besides, by monitoring a signalcontinuously and examining the loci of the signal over time, it ispossible to obtain an unlimited measurable phase range beyond the rangefrom −180° to 180°.

In order to achieve such a widening of the measurable phase range, it isdesirable to dispose the analyte detecting portion 111 which ismonotonically increased in mass in response to analyte detection, aswell as to carry out measurement signal sampling two or more times attime-spaced intervals. The measurement interval is determined inaccordance with a rate at which a reaction to induce mass changesproceeds. It is more desirable to carry out measurement signal samplingconsecutively.

As described hereinabove, it is possible to provide an analyte sensor100 which is capable of detection with fewer constituent components andwith fewer signal processing steps in a measurable phase range as wideas that of the quadrature modulation system.

Structure of Analyte Sensor 100A

Next, referring to FIG. 2, the structure of an analyte sensor 100A whichis an embodiment of the principle of the analyte sensor 100 will bedescribed.

As shown in FIG. 2 which is a perspective view, from the standpoint ofappearance, the analyte sensor 100A is composed mainly of apiezoelectric substrate 1 and a cover 3. The cover 3 is formed with afirst through hole 18 acting as an inlet for an analyte solution, and anair slot or a second through hole 19 acting as an outlet for an analytesolution.

FIG. 3 shows a perspective view of the SAW sensor 100A, with one-half ofthe cover 3 removed. As shown in the drawing, a space 20 acting as aflow path for an analyte (solution) is formed inside the cover 3. Thefirst through hole 18 is in communication with the space 20. That is, ananalyte admitted from the first through hole 18 flows into the space 20.

The analyte solution which has flowed into the space 20 contains atarget substance which is an analyte, and, the analyte reacts with adetecting portion made of a metal film 7 and so forth formed on thepiezoelectric substrate.

The piezoelectric substrate 1 is formed of a piezoelectricsingle-crystal substrate, such for example as a lithium tantalate(LiTaO₃) single crystal, a lithium niobate (LiNbO₃) single crystal, or aquartz. The planar shape and dimensions of the piezoelectric substrate 1may be determined arbitrarily. As an example, the piezoelectricsubstrate 1 has a thickness of 0.3 mm to 1 mm.

FIGS. 4A and 4B show sectional views of the SAW sensor 100. FIG. 4A is asectional view taken along the line IVa-IVa shown in FIG. 2, and FIG. 4Bis a sectional view taken along the line IVb-IVb shown in FIG. 2.Moreover, FIG. 5 shows a top view of the SAW sensor 100, with the cover3 removed.

As shown in FIGS. 4B and 5, a first IDT electrode 5 a, a second IDTelectrode 6 a, a reference first IDT electrode 5 b, and a referencesecond IDT electrode 6 b are formed on the upper surface of thepiezoelectric substrate 1. The first IDT electrode 5 a and the referencefirst IDT electrode 5 b are intended for generation of predeterminedSAW, and the second IDT electrode 6 a and the reference second IDTelectrode 6 b are intended to receive SAW generated by the first IDTelectrode 5 a and SAW generated by the reference first IDT electrode 5b, respectively. In order for the second IDT electrode 6 a to receiveSAW generated by the first IDT electrode 5 a, the second IDT electrodeis placed on a propagation path of SAW generated by the first IDTelectrode. The reference second electrode 6 b is placed similarly inrelation to the reference first IDT electrode 5 b.

Since the reference first IDT electrode 5 b and the reference secondelectrode 6 b are similar to the first IDT electrode 5 a and the secondIDT electrode 6 a, respectively, in what follows, the first IDTelectrode 5 a and the second IDT electrode 6 a will be quoted by way ofillustration.

The first IDT electrode 5 a and the second IDT electrode 6 a comprise apair of comb-like electrodes (refer to FIG. 5). Each of the comb-likeelectrodes includes two bus bars opposed to each other and a pluralityof electrode fingers extending from one of the bus bars toward theother. The comb-like electrode pair is placed so that the plurality ofelectrode fingers are arranged in an interdigitated pattern. The firstIDT electrode 5 a and the second IDT electrode 6 a constitute atransversal-type IDT electrode.

Frequency characteristics can be designed on the basis of the number ofthe electrode fingers of the first IDT electrode 5 a and the second IDTelectrode 6 a, the distance between the adjacent electrode fingers, thecrossing width of the electrode fingers, etc. used as parameters. Thetypes of SAW excited by the IDT electrode include: Rayleigh wave; Lovewave; and Leaky wave, and, in the detection element 3, Love wave isutilized.

An elastic member for suppressing SAW reflection may be provided in aregion outside of the first IDT electrode 5 a in the propagationdirection of SAW. The frequency of SAW can be set within a range of fromseveral megahertz (MHz) to several gigahertz (GHz), for example.Particularly, it is advisable to set the SAW frequency within a range offrom several hundred MHz to 2 GHz as a matter of practicality, and also,this makes it possible to achieve downsizing of the piezoelectricsubstrate 1 with a consequent miniaturization of the SAW sensor 100A.

The first IDT electrode 5 a and the second IDT electrode 6 a are eachconnected to a pad 9 via a wiring line 8. A signal is inputted from theoutside to the first IDT electrode 5 a through the pad 9 and the wiringline 8, and is outputted to the outside from the second IDT electrode 6a. As shown in FIG. 5, by separately providing a pad for the signalinput side and a pad for the signal output side, on the opposite sides,respectively, of the piezoelectric substrate 1, it is possible to lessenthe influence of a signal from the input side and a signal from theoutput side.

A short-circuit electrode 10 a is formed in a first region 1 a which isa region of the upper surface of the piezoelectric substrate 1 which islocated between the first IDT electrode 5 a and the second IDT electrode6 a. The short-circuit electrode 10 a is intended to cause electricalshort-circuiting in a part of the upper surface of the piezoelectricsubstrate 1 which serves as the SAW propagation path. The provision ofthe short-circuit electrode 10 a makes it possible to, depending on thetype of SAW, reduce SAW losses. It can be considered that theloss-reduction effect brought about by the short-circuit electrode 10 abecomes especially high when Leaky wave is used as SAW.

For example, the short-circuit electrode 10 a has a rectangular shapeelongated along the SAW propagation path from the first IDT electrode 5a toward the second IDT electrode 6 a. The width of the short-circuitelectrode 10 a in a direction perpendicular to the SAW propagationdirection (x direction) is equal to, for example, the crossing width ofthe electrode fingers of the first IDT electrode 5 a. Moreover, thefirst IDT electrode-sided end of the short-circuit electrode 10 a in adirection parallel to the SAW propagation direction (y direction) isspaced away from the center of the electrode finger situated at the endof the first IDT electrode 5 a by a distance equal to one-halfwavelength of SAW. Likewise, the second IDT electrode-sided end of theshort-circuit electrode 10 a in the y direction is spaced away from thecenter of the electrode finger situated at the end of the second IDTelectrode 6 a by a distance equal to one-half wavelength of SAW.

The short-circuit electrode 10 a may be in an electrically floatingstate, or alternatively, a pad 9 for ground potential may be provided,and the short-circuit electrode 10 a may be connected to this pad 9 soas to stand at a ground potential. In the case where the short-circuitelectrode 10 a is set to the ground potential, it is possible tosuppress propagation of a direct wave resulting from electromagneticcoupling between the first IDT electrode 5 a and the second IDTelectrode 6 a.

Likewise, a short-circuit electrode 10 b is formed in a first region 1 bwhich is a region between two electrodes, namely the reference first IDTelectrode 5 b and the reference second IDT electrode 6 b.

The first IDT electrode 5 a, the second IDT electrode 6 a, the referencefirst IDT electrode 5 b, the reference second IDT electrode 6 b, theshort-circuit electrodes 10 a and 10 b, the wiring line 8, and the pad 9are made of aluminum, an alloy of aluminum and copper, or the like, forexample. Moreover, the electrodes may be given a multilayer structure.In the case of adopting the multilayer structure, for example, the firstlayer is made of titanium or chromium, and the second layer is made ofaluminum or an aluminum alloy.

The first IDT electrode 5 a, the second IDT electrode 6 a, the referencefirst IDT electrode 5 b, the reference second IDT electrode 6 b, and theshort-circuit electrodes 10 a and 10 b are covered with a protectivefilm 4. The protective film 4 contributes to the protection of each ofthe electrodes and the wiring from oxidation, for example. Theprotective film 4 is made of silicon oxide, aluminum oxide, zinc oxide,titanium oxide, silicon nitride, silicon, or the like. In the SAW sensor100A, silicon dioxide (SiO₂) is used for the protective film 4.

The protective film 4 is formed over the entire upper surface of thepiezoelectric substrate 1, with the pads 9 exposed. The first IDTelectrode 5 a and the second IDT electrode 6 a are covered with theprotective film 4. This makes it possible to suppress the corrosion ofthe IDT electrodes.

The protective film 4 has a thickness of 100 nm to 10 μm, for example.The protective film 4 does not necessarily have to be formed over theentire upper surface of the piezoelectric substrate 1, and therefore,for example, it may be formed so as to cover only the central area ofthe upper surface of the piezoelectric substrate 1, with the other areaalong the outer periphery of the upper surface of the piezoelectricsubstrate 1 including the pads 9 exposed.

As shown in FIG. 4A, the first IDT electrode 5 a is accommodated in afirst vibration space 11 a, and the second IDT electrode 6 a isaccommodated in a second vibration space 12 a. This makes it possible toseparate the first IDT electrode 5 a and the second IDT electrode 6 afrom outside air and an analyte solution, and thereby protect the firstIDT electrode 5 a and the second IDT electrode 6 a against acorrosion-inducing substance such as water. Moreover, by securing thefirst vibration space 11 a and the second vibration space 12 a, thefirst IDT electrode 5 a and the second IDT electrode 6 a can be kept ina condition where SAW excitation will not be seriously hindered.

The first vibration space 11 a and the second vibration space 12 a canbe formed by joining a plate-like body 2 having recesses for creatingthese vibration spaces to the piezoelectric substrate 1.

Likewise, a first vibration space 11 b and a second vibration space 12 bare secured for the reference first IDT electrode 5 b and the referencesecond IDT electrode 6 b.

Although the first vibration space 11 and the second vibration space 12of the SAW sensor 100A are each a rectangular parallelepiped space, theshape of the vibration space is not limited to the rectangularparallelepiped shape, and can therefore be changed to another shape onan as needed basis with consideration given to the form or arrangementof the IDT electrodes, and more specifically, for example, the vibrationspace may be dome-shaped as seen in a sectional view, or may beelliptically shaped as seen in a plan view.

The plate-like body 2 has, in a region between the recesses for creatingthe first vibration space 11 a and the second vibration space 12 a, athrough part formed therethrough in a thickness direction thereof. Thisthrough part is intended for the formation of a metal film 7 a on theSAW propagation path. That is, when the plate-like body 2 joined to thepiezoelectric substrate 1 is seen in a plan view, at least part of thepropagation path of SAW propagating from the first IDT electrode 5 a tothe second IDT electrode 6 a is exposed from the through part, and themetal film 7 a is formed on this exposed part.

Likewise, the plate-like body 2 also has, in a region between therecesses for creating the first vibration space 11 b and the secondvibration space 12 b, another through part formed therethrough in thethickness direction thereof. This through part is intended for theformation of a metal film 7 b on the SAW propagation path.

The plate-like body 2 having such a shape can be formed with use of aphotosensitive resist, for example.

The metal film 7 a exposed from the through part of the plate-like body2 constitutes an analyte detecting portion for an analyte solution. Themetal film 7 a has a double-layer structure consisting of titanium orchromium and gold laminated thereon, for example. An aptamer such forexample as a nucleic acid- or peptide-made aptamer is immobilized on thesurface of the metal film 7 a. Upon occurrence of contact between ananalyte solution and the aptamer-immobilized metal film 7 a, a specifictarget substance contained in the analyte solution is bound to theaptamer adaptable to the target substance. In such a structure, theanalyte is bound to the aptamer, and, as adsorption proceeds, the massof the metal film 7 a is monotonically increased. That is, there arisesa monotonic increase in mass in response to analyte detection. Note thatthe mass of the metal film 7 a is monotonically increased only duringthe interval when the analyte is being continuously supplied onto themetal film 7 a. For example, in a case where a buffer solution issupplied subsequent to the supply of the analyte before and after thesupply of the analyte solution, even if the analyte passes over themetal film 7 a and the mass is reduced by the separation between theanalyte and the aptamer, there is no problem.

Moreover, the metal film 7 b exposed from the other through part of theplate-like body 2 constitutes a reference measuring portion. The metalfilm 7 b has a double-layer structure consisting of titanium or chromiumand gold laminated thereon, for example. The surface of the metal film 7b is free from aptamer immobilization so as not to exhibit reactivity tothe analyte. Instead, the metal film may be subjected to surfacetreatment to cause reduced response to the analyte solution forstabilizing purposes.

In order to assess the properties and so forth of the analyte solutionthrough the use of SAW, to begin with, a predetermined voltage (signal)is applied, through the pad 9 and the wiring line 8, to the first IDTelectrode 5 a from external measurement equipment. Then, the surface ofthe piezoelectric substrate 1 is excited within the region where thefirst IDT electrode 5 a is formed, thereby producing SAW having apredetermined frequency. Part of the thusly produced SAW passes throughthe first region 1 a which is the region between the first IDT electrode5 a and the second IDT electrode, and then reaches the second IDTelectrode 6 a. At this time, in the metal film 7 a situated on the firstregion 1 a, the aptamer immobilized on the metal film 7 a is bound tothe specific target substance contained in the analyte solution, and theweight of the metal film 7 changes by a weight corresponding to thebound amount, which results in variations in the phase characteristicsand so forth of SAW passing under the metal film 7 a. Upon the SAW whichhas undergone characteristics variations reaching the second IDTelectrode 6 a, a corresponding voltage is developed in the second IDTelectrode 6 a. This voltage is outputted, through the wiring line 8 andthe pad 9, to the outside as a detection signal in the form of an ACsignal. The properties and ingredients of the analyte solution can beexamined by processing the signal in the mixer 130 as shown in FIG. 1.

That is, the piezoelectric substrate 1, the metal film 7 a acting as theanalyte detecting portion formed on the piezoelectric substrate 1, thefirst IDT electrode 5 a, and the second IDT electrode 6 a constitute adetection element 110A.

Likewise, in the same space 20, the other metal film 7 b free fromaptamer immobilization is disposed, and, an AC signal outputted from thereference second IDT electrode 6 b in the wake of inputting of a signalfrom the reference first IDT electrode 5 b is defined as a referencesignal for use in calibration of signal fluctuations caused byenvironmental variations such as variations in temperaturecharacteristics and humidity.

That is, the piezoelectric substrate 1, the metal film 7 b acting as thereference measuring portion formed on the piezoelectric substrate 1, thereference first IDT electrode 5 b, and the reference second IDTelectrode 6 b constitute a reference element 120A.

In the case of conducting measurement through the use of SAW in thatway, as has already been described, there is a need to provide aprotective film such as silicon oxide to protect the IDT electrodes andso forth, but, as the result of a survey by the inventors of the presentapplication, it has been found that, if such a protective film isexposed inside the flow path for an analyte solution, a trouble such asgreat variations of the detection sensitivity or deterioration of thedetection sensitivity is easy to occur.

Although the cause of such a trouble has not been clarified, this isprobably attributable to a phenomenon in which an aptamer adheres to theprotective film 4 exposed from the through part when the aptamer isimmobilized on the metal film 7 a and consequently a desired amount ofthe aptamer cannot be immobilized on the metal film 7 a, or a targetsubstance (analyte) adheres to the protective film 4 when an analytesolution is charged into the space 20.

In light of this, the SAW sensor 100A is designed so that the protectivefilm 4 is not exposed inside the space 20 acting as a flow path.

In the interest of uniformity in the amount of an analyte solutionduring measurement, in the SAW sensor 100A is provided the space 20acting as a flow path for an analyte solution. The space 20 of the SAWsensor 100A is a space surrounded with the inner surface of the cover 3,the outer surface of the plate-like body 2, and the upper surface of themetal film 7 a, 7 b.

Since such a space 20 has basically a constant volumetric capacity, bycharging an analyte solution into the space 20, it is possible to renderthe amount of the analyte solution uniform during measurement.

In charging an analyte into the space 20, a capillary phenomenon isexploited in the SAW sensor 100A. Specifically, by adjusting each of thesize (diameter, for example) of the first through hole 18 acting as ananalyte inlet and the size (width and height, for example) of the space20 acting as the flow path for an analyte solution to a predeterminedvalue in consideration of the type of the analyte solution, the materialused for the cover 3, etc., it is possible to urge an analyte to movefrom the inlet to the flow path and eventually to the analyte detectingportion by exploiting the capillary phenomenon. The width w (FIG. 4A) ofthe space 20 falls in a range of from 0.5 mm to 3 mm, for example, and,the height h (FIG. 4A) thereof falls in a range of from 0.05 mm to 0.5mm, for example. The diameter of the first through hole 18 falls in arange of from 50 μm to 500 μm, for example.

With the formation of the first through hole 18 and the space 20, simplyby bringing an analyte into contact with the opening of the firstthrough hole 18, the analyte can automatically be drawn into the space20 by capillary action, and the space 20 is filled with the analyte.Thus, according to the SAW sensor 100, the SAW sensor in itself includesan analyte-solution suction mechanism, wherefore the suction of ananalyte can be effected without the necessity of using an instrumentsuch as a pipette. Note that the shape of the first through hole 18acting as the analyte inlet is not limited to a cylindrical shape, andtherefore, for example, the first through hole 18 may be so shaped thatits diameter becomes smaller or larger gradually toward the space 20, ormay have a rectangular opening. Moreover, the forming position of thefirst through hole 18 is not limited to the ceiling part of the cover 3,but may be in a side wall of the cover 3.

In addition to the first through hole 18, the second through hole 19 isformed in the cover 3. The second through hole 19 is located at an endpart of the cover opposite to the end part bearing the first throughhole 18, and is in communication with the space 20. By virtue of such asecond through hole 19, when an analyte enters the space 20, air whichis originally present in the space 20 is expelled to the outside fromthe second through hole 19, whereby the analyte can be easily admittedinto the space 20.

The corners of that part of the space 20 which is defined by the innersurface of the cover 3 are rounded off. For example, as shown in thesectional view of FIG. 4, the juncture of the first through hole 18 andthe space 20, the juncture of the second through hole 19 and the space20, and the juncture of the inner periphery of the cover 3 and theplate-like body 2 are each rounded off.

If the corners of the space 20 acting as the flow path for an analytesolution become angular, an analyte solution will accumulate at thecorner, and consequently the analyte tends to be stagnant. In thepresence of analyte stagnation, for example, there arise concentrationvariations within the target substance of the analyte charged in thespace 20, which gives rise to a problem such as detection sensitivitydeterioration. By contrast, where the space 20 has rounded corners as inthe analyte sensor 100A, the analyte is less prone to stagnation,whereby the concentration of the target substance can be rendereduniform throughout the space 20.

Moreover, it is advisable to form the first through hole 18 acting asthe analyte inlet in a position as close to the end of the space 20 aspossible from the viewpoint of preventing analyte accumulation.

For example, the cover 3 is made of polydimethylsiloxane. With use ofpolydimethylsiloxane as the material for the cover 3, the cover 3 can begiven a desired shape, for example, the cover 3 can be configured tohave rounded corners. Moreover, with use of polydimethylsiloxane, theceiling part and the side wall of the cover 3 can be made thickrelatively easily. For example, the ceiling part and the side wall ofthe cover 3 have a thickness of 1 mm to 5 mm.

In the analyte sensor 100A, the cover 3 is disposed, with the outerperiphery of its lower surface kept in contact with the protective film4 situated around the plate-like body 2, so as to be joined to theprotective film 4 at the contacted part. In other words, the cover 3 canbe deemed to be joined to the piezoelectric substrate 1 through theprotective film 4. In a case where the cover 3 is made ofpolydimethylsiloxane, and the protective film 4 is made of SiO₂, byperforming oxygen plasma treatment on the surface of the cover 3 to becontacted by the protective film 4, it is possible to join the cover 3directly to the protective film 4 without the necessity of using anadhesive or the like. Although the reason why the cover 3 and theprotective film 4 can be directly joined to each other under such acondition has not been clarified, this is probably because a covalentbond of Si and O is formed between the cover 3 and the protective film4.

As has already been described, the substrate is shared between thedetection element 110A and the reference element 120A. In such astructure, there is the possibility of occurrence of crosstalk betweensignals on both elements. Therefore, as shown in FIG. 5, a referencepotential line 31 connected to a reference potential is disposed betweena region serving as the detection element 110A indicated by broken linesin the drawing and a region serving as the reference element 120Aindicated by dotted lines in the drawing on the piezoelectric substrate1. By virtue of the reference potential line 31, occurrence of crosstalkbetween the detection element 110A and the reference element 120A can beprevented, wherefore a high-sensitivity analyte sensor 100A can beprovided.

As shown in FIG. 5, the reference potential line 31 is connected withone of the paired comb-like electrodes constituting each of the firstIDT electrode 5 a, the second IDT electrode 6 a, the reference first IDTelectrode 5 b, and the reference second IDT electrode 6 b. Out of thepaired comb-like electrodes constituting each of the first IDT electrode5 a, the second IDT electrode 6 a, the reference first IDT electrode 5b, and the reference second IDT electrode 6 b, the electrode to beconnected to the reference potential is located at the side on which thereference potential line 31 is disposed. In other words, of the pairedcomb-like electrodes, the inwardly located electrode is connected to thereference potential.

Such an arrangement makes it possible to facilitate the layout of thewiring lines 8 for the detection element 110A and the reference element120A, as well as to render the wiring lines 8 uniform in length. Thus,the reference signal from the reference element 120A becomes a moreaccurate signal for reference purposes.

Moreover, in a case where the structure including the detection element110A and the reference element 120A and the structure including themeasurement portion 140 and the detection amount calculation portion 150are provided independently of each other, in the structure including themeasurement portion 140 and the detection amount calculation portion150, it is desirable to conduct wiring installation in conformity withthe arrangement of the wiring lines 8 on the piezoelectric substrate 1.This configuration makes it possible to prevent crosstalk even in thestructure including the measurement portion 140 and the detection amountcalculation portion 150.

Modified Example: Arrangement of Analyte Flow Path, Detection Element,and Reference Element

Although in the above embodiment it is described that the constituentcomponents are so arranged that the direction of elongation of the space20 acting as the analyte flow path and the propagation direction ofsurface acoustic waves in the detection element 110A and the referenceelement 120A are perpendicular to each other, as in an analyte sensor100B as shown in FIGS. 6A and 6B, the directions may be parallel to eachother.

Modified Example: Arrangement of Analyte Flow Path, Detection Element,and Reference Element

Although in the above embodiment it is described that the space 20acting as the analyte flow path is shared between the detection element110A and the reference element 120A, as in an analyte sensor 100C asshown in FIGS. 7A and 7B, the space 20 may be provided specifically foreach of the detection element 110A and the reference element 120A.While, in the example shown in FIGS. 7A and 7B, the inlet 18 is providedfor each space 20 on an individual basis, the inlet can be sharedbetween two analyte flow paths. Moreover, while, in the example shown inFIGS. 7A and 7B, like the example shown in FIGS. 6A and 6B, the SAWpropagation direction and the analyte-flow-path elongation directioncoincide with each other, the SAW propagation direction and theanalyte-flow-path elongation direction may be perpendicular to eachother.

Modified Example: Piezoelectric Substrate 1

Although in the above embodiment it is described that a piezoelectricsubstrate is shared between the detection element 110A and the referenceelement 120A, an element substrate for the detection element 110A and areference element substrate for the reference element 120A may beprovided independently of each other. In this case, occurrence ofcrosstalk between the detection element 110A and the reference element120A can be prevented without fail. Moreover, in this case, it isadvisable to provide two separate base bodies that hold the elementsubstrate and the reference element substrate, respectively.

Modified Example: π/2 Delay Line

Although in the above embodiment it is described that the first IDTelectrode 5 a, the second IDT electrode 6 a, the reference first IDTelectrode 5 b, and the reference second IDT electrode 6 b are connectedto their nearby pads 9 without taking any detour, as in an analytesensor 100D as shown in FIG. 8, a π/2 delay line 32 may be disposedbetween the pad and, one of the paired comb-like electrodes constitutingthe second IDT electrode 6 a, the one comb-like electrode being notconnected to the reference potential. In the analyte sensor 100A, sincesignal processing operation is performed by the heterodyne system, itfollows that a signal makes a sine curve. Therefore, the slope of thesine curve with respect to phases corresponding to 0° and ±180°decreases, which results in a decline in sensitivity. However, in theanalyte sensor, the vicinity of 0° generally corresponds to a rise of asignal change entailed by analyte detection, wherefore it is desiredthat this range should be measured with high sensitivity. In light ofthis, by providing the π/2 delay line 32 for one of the constituentelectrodes of the second IDT electrode 6 a to be connected to the mixer,it is possible to change the signal phase and thereby achieveenhancement in sensitivity in the vicinity of 0°.

Such a π/2 delay line 32 can be prepared by forming a conductor film onthe piezoelectric substrate 1, and then performing patterning thereon soas to obtain a necessary line length.

Moreover, as shown in FIG. 8, by arranging the pads 9 side by side atthe edge of one side constituting the piezoelectric substrate 1, thehandling becomes easier, and also the wiring lines 8 can be laid outwith consistency, wherefore wiring-induced signal delay, signal shift,and noise superimposition can be suppressed.

Another Embodiment

In the above embodiment it is described that signals from the detectionelement 110 and the reference element 120 are synthesized directly bythe mixer 130. By contrast, in an analyte sensor 100E as shown in FIG.9, low-noise amplifiers 132 are disposed between the detection element110 and the mixer 130, and between the reference element 120 and themixer 130, respectively. That is, a measurement portion 140E includesthe low-noise amplifiers 132, the mixer 130, and the low-pass filter131.

In the SAW sensor, in general, high sensitivity can lead to significantvariations in amplitude characteristics. That is, when the SAW sensor isdesigned to have high sensitivity by making adjustment to the thicknessof the protective film 4 and so forth, a large loss may occur, whichcauses the possibility of a failure of accurate measurement. On theother hand, when a signal inputted to the mixer 130 is small, noise maybe increased, which causes the possibility of impairment of detectionaccuracy.

Furthermore, when signals inputted to the detection element 110 and thereference element 120 are large, there is the possibility of crosstalkbetween output signals on the detection element 110 and the referenceelement 120. In addition, when signals inputted to the detection element110 and the reference element 120 are large, there is the possibilitythat output signals from the detection element 110 and the referenceelement 120 will leak to the outside as electromagnetic waves.

It will thus be seen that the placement of the low-noise amplifiers 132between the detection element 110 and the mixer 130 and between thereference element 120 and the mixer 130, is important for the attainmentof high detection accuracy.

Examples

One and the same analyte was measured by each of the analyte sensor 100Eand the analyte sensor 100 devoid of the low-noise amplifiers 132.Specifically, the measurement was conducted under conditions where: thecenter frequency of SAW is 414 MHz; the IDT electrodes 5 and 6 are madeof Al and are 300 nm in thickness; the protective film 4 is made of SiO₂and is 100 nm in thickness; and the distance between the first IDTelectrode 5 a and the IDT electrode 6 a is 300λ (λ represents thewavelength of SAW propagating through the metal film 7 after beingexcited by the first IDT electrode 5 a). Moreover, there were preparedanalytes that have contained target concentrations of 100 nM, 200 nM,and 500 nM, respectively, and these analytes were supplied to theanalyte sensors. The measurement was also conducted by means of a vectornetwork analyzer (VNA) as a reference example.

FIG. 11 shows the result of actual measurement of the values of changesin phase with respect to time since the supply of analyte solutions tothe analyte sensors 100 and 100E. In FIG. 11, the amount of phase change(θ) is taken along the ordinate axis, and time (sec) is taken along theabscissa axis. The solid explanatory legends indicate the result ofmeasurement by the LNA-free analyte sensor 100, whereas the hollowexplanatory legends indicate the result of measurement by theLNA-equipped analyte sensor 100E. As shown in FIG. 11, it can beconfirmed that measurement has been conducted properly for a long periodof time. That is, the analyte sensor pursuant to the invention can beconfirmed to be capable of measurement in an even wider phase range bythe heterodyne system. Also, the dependence of the analyte solution onconcentration can be confirmed.

Moreover, the analyte sensor 100E equipped with the low-noise amplifiers132 was found to be far smaller than the analyte sensor devoid of thelow-noise amplifiers 132 in respect of variations in phase change value.Specifically, as shown in FIG. 12, the analyte sensor 100E comparesfavorably in measurement accuracy with VNA which is capable of low-levelmeasurement and is also able to increase the magnitude of an inputsignal in itself. Note that FIG. 12 is a chart showing the result ofmeasurement by VNA and the result of measurement by the analyte sensor100E.

Method for Measuring Detection Amount of Analyte

An analyte sensing method adopted in the analyte sensor will bedescribed.

Analyte-Solution Supplying Step

The first step is an analyte-solution supplying step of supplying ananalyte containing a target to the analyte detecting portion of thedetection element that is increased in mass in response to targetadsorption or reaction with the target, and the reference detectingportion of the reference element that undergoes neither targetadsorption nor reaction with the target.

Determination Step

Next, in accordance with the heterodyne system, a measurement signal isobtained from a detection signal which is an AC signal responsive tomass variations in the analyte detecting portion and a reference signalfrom the reference detecting portion that is an AC signal relative tothe detection signal.

Then, as a phase-change value determination step, two candidatephase-change values of a positive value and a negative value, arederived from the measurement signal, and whether the phase is positiveor negative is judged on the basis of temporal changes in measurementsignal strength, so that a phase change value can be determined from thetwo candidate phase-change values. In this example, since themeasurement signal is processed by the heterodyne system, it followsthat there are two candidate phase-change values, wherefore phasechanges cannot be directly ascertained from signal strength. Note thatthe analyte detecting portion is so designed that the mass is increasedin response to target detection, wherefore there is a monotonic increasein phase change. In this regard, by adding a step of examining temporalchanges in measurement signal strength, it is possible to identifywhether the strength is on the increase or on the decrease, and therebymake a decision about which one of the two candidate phase-change valuesto be selected. Specifically, when the measurement signal strength isdecreased with time, the positive value is determined as the phasechange value, and when the measurement signal strength is increased withtime, the negative value is determined as the phase change value.

In order to accomplish such steps, specifically, it is advisable toconduct measurement of mixed signal strength at least twice attime-spaced intervals. It is also advisable to carry out measurementsignal sampling consecutively.

Calculation Step

Lastly, a calculation step of calculating the detection amount of ananalyte on the basis of the thusly determined phase change value, isperformed.

By following the above-described procedure, the detection amount of ananalyte can be measured.

Moreover, in the determination step, each of the detection signal andthe reference signal may be amplified, and, in this case, a measurementsignal is obtained on the basis of these detection and reference signalsamplified by the heterodyne system.

The invention is not limited to the embodiments as describedhereinabove, and may therefore be carried out in various forms. Forexample, although the analyte sensor 100A is illustrated as beingdesigned so that the detecting portion comprises the metal film 7 andthe aptamer immobilized on the surface of the metal film 7, in a casewhere the target substance contained in the analyte solution reacts withthe metal film 7, the detecting portion may be composed solely of themetal film 7 without using an aptamer.

Although in the above embodiment it is described that the analytedetecting portion is monotonically increased in mass in response toadsorption of a target contained in an analyte or reaction with thetarget, it is possible to employ an analyte detecting portion which ismonotonically decreased in mass in response to reaction with a targetcontained in an analyte. In this case, for example, the analytedetecting portion can be implemented by immobilizing, on a Au film, areactive group which exhibits reactivity to the target and has aconformation in which part of the reactive group comes off through thereaction with the target. Then, a choice between the condition (2) andthe condition (4) is made for phase-change value calculation based onthe heterodyne system. For example, under the condition (4), when themeasurement signal strength is increased with time, the positivecandidate phase-change value is assigned, and when the measurementsignal strength is decreased with time, the negative candidatephase-change value is assigned. Such a configuration affordsadaptability to an analyte detecting portion of a type which ismonotonically changed in mass in response to adsorption of a targetcontained in an analyte or reaction with the target.

REFERENCE SIGNS LIST

-   -   1: Piezoelectric substrate    -   2: Plate-like body    -   3: Cover    -   4: Protective film    -   5 a: First IDT electrode    -   5 b: Reference first IDT electrode    -   6 a: Second IDT electrode    -   6 b: Reference second IDT electrode    -   7 a, 7 b: Metal film    -   8: Wiring line    -   9: Pad    -   10: Short-circuit electrode    -   11 a, 11 b: First vibration space    -   12 a, 12 b: Second vibration space    -   20: Space    -   31: Reference potential line    -   32: π/2 delay line

The invention claimed is:
 1. An analyte sensor, comprising: a detection signal output portion which outputs a detection signal which changes in response to adsorption of a target provided in an analyte or to reaction with the target; a reference signal output portion which outputs a reference signal which serves as a reference signal relative to the detection signal; a measurement portion configured to determine two candidate phase-change values from a measurement signal which is obtained based on the detection signal and the reference signal in accordance with a heterodyne system, and output one of the two candidate phase-change values as a phase change value based on temporal changes in the measurement signal, wherein the measurement portion comprises a mixer configured to receive and mix the measurement signal and a low-pass filter configured to receive the mixed measurement signal from the mixer and filter out higher-frequency components of the mixed measurement signal, wherein a first phase-change value from among the two candidate phase-change values is outputted when a strength of the measurement signal is increasing and a second phase-change value from among the two candidate phase-change values is outputted when the strength of the measurement signal is decreasing; and a detection amount calculation portion wherein an amount of the target in the analyte is calculated based on the output of the first phase-change value or the second phase-change value.
 2. The analyte sensor according to claim 1, wherein the temporal changes in measurement signal are obtained by conducting measurement two or more times at time-spaced intervals.
 3. The analyte sensor according to claim 1, wherein the temporal changes in measurement signal are obtained by conducting continuous measurement.
 4. The analyte sensor according to claim 1, further comprising a detection amount calculation portion which calculates a detection amount of the analyte on a basis of the phase change value.
 5. The analyte sensor according to claim 1, wherein the detection signal output portion comprises an analyte detecting portion which changes in response to adsorption of the target or to reaction with the target.
 6. The analyte sensor according to claim 5, wherein the measurement portion is configured to output a larger one of the two candidate phase-change values as the phase change value when the temporal changes in measurement signal decrease with time, and output a smaller one of the two candidate phase-change values as the phase change value when the temporal changes in measurement signal increase with time, in a case where a mass of the analyte detecting portion increases and the measurement signal is obtained by subtracting the detection signal from the reference signal, or where the mass of the analyte detecting portion decreases and the measurement signal is obtained by subtracting the reference signal from the detection signal, and the measurement portion is configured to output a smaller one of the two candidate phase-change values as the phase change value when the temporal changes in measurement signal decrease with time, and output a larger one of the two candidate phase-change values as the phase change value when the temporal changes in measurement signal increase with time, in a case where the mass of the analyte detecting portion increases and the measurement signal is obtained by subtracting the reference signal from the detection signal, or where the mass of the analyte detecting portion decreases and the measurement signal is obtained by subtracting the detection signal from the reference signal.
 7. The analyte sensor according to claim 1, wherein the two candidate phase-change values are composed of a positive candidate phase-change value and a negative candidate phase-change value.
 8. The analyte sensor according to claim 1, further comprising an analyte flow path through which the analyte can be supplied to the detection signal output portion and the reference signal output portion in order or simultaneously.
 9. The analyte sensor according to claim 8, wherein the detection signal output portion comprises an analyte detecting portion which changes in response to adsorption of the target or to reaction with the target, and the reference signal output portion comprises a reference measuring portion which undergoes neither adsorption of the target nor reaction with the target, and through the analyte flow path, the analyte can be supplied to the analyte detecting portion and the reference measuring portion in order or simultaneously.
 10. The analyte sensor according to claim 1, wherein the reference signal output portion comprises a reference measuring portion which undergoes neither adsorption of the target nor reaction with the target.
 11. The analyte sensor according to claim 5, wherein the detection signal output portion comprises a piezoelectric detection element substrate, the analyte detecting portion is placed on the piezoelectric detection element substrate, and the detection signal output portion comprises a detection first IDT electrode which is placed on the piezoelectric detection element substrate and is configured to produce a first elastic wave toward the analyte detecting portion, and a detection second IDT electrode which is placed on the piezoelectric detection element substrate and is configured to receive the first elastic wave which has passed through the analyte detecting portion.
 12. The analyte sensor according to claim 11, wherein the detection signal is an AC signal obtained when the first elastic wave which has passed through the analyte detecting portion is received by the detection second IDT electrode.
 13. The analyte sensor according to claim 10, wherein the reference signal output portion comprises a piezoelectric reference element substrate, the reference measuring portion is placed on the piezoelectric reference element substrate, and the reference signal output portion comprises a reference first IDT electrode which is placed on the piezoelectric reference element substrate and is configured to produce a second elastic wave toward the reference measuring portion, and a reference second IDT electrode which is placed on the piezoelectric reference element substrate and is configured to receive the second elastic wave which has passed through the reference measuring portion.
 14. The analyte sensor according to claim 13, wherein the reference signal is an AC signal obtained when the second elastic wave which has passed through the reference measuring portion is received by the reference second IDT electrode.
 15. The analyte sensor according to claim 11, wherein the reference signal output portion comprises a reference measuring portion which undergoes neither adsorption of the target nor reaction with the target, and the reference signal output portion comprises a piezoelectric reference element substrate, the reference measuring portion is placed on the piezoelectric reference element substrate, and the reference signal output portion comprises a reference first IDT electrode which is placed on the piezoelectric reference element substrate and is configured to produce a second elastic wave toward the reference measuring portion, and a reference second IDT electrode which is placed on the piezoelectric reference element substrate and is configured to receive the second elastic wave which has passed through the reference measuring portion.
 16. The analyte sensor according to claim 15, wherein the detection element substrate and the reference element substrate are formed integrally with each other, and further comprising a reference potential line located between a detection element region where the analyte detecting portion, the detection first IDT electrode, and the detection second IDT electrode are disposed, and a reference element region where the reference measuring portion, the reference first IDT electrode, and the reference second IDT electrode are disposed.
 17. The analyte sensor according to claim 16, wherein the detection first IDT electrode, the detection second IDT electrode, the reference first IDT electrode, and the reference second IDT electrode are each composed of a pair of comb-like electrodes, and one of the pair of comb-like electrodes is connected to the reference potential line.
 18. The analyte sensor according to claim 16, further comprising an analyte flow path through which the analyte can be supplied to the detection signal output portion and the reference signal output portion in order or simultaneously, wherein a direction of elongation of the analyte flow path is parallel to a propagation direction of the first elastic wave and the second elastic wave.
 19. The analyte sensor according to claim 1, further comprising: a π/2 delay line disposed between the detection signal output portion and the measurement portion, and/or between the reference signal output portion and the measurement portion, the π/2 delay line being configured to permit passage of the detection signal or the reference signal which is prior to acquisition of the measurement signal by the heterodyne system.
 20. The analyte sensor according to claim 1, further comprising: low-noise amplifiers which are disposed between the detection signal output portion and the measurement portion, and between the reference signal output portion and the measurement portion, respectively, the low-noise amplifiers being configured to amplify the detection signal and the reference signal, respectively.
 21. An analyte sensing method, comprising: an analyte supplying step of supplying an analyte in which a target is provided, to a detection signal output portion which outputs a detection signal which changes in response to adsorption of the target or to reaction with the target, and a reference signal output portion which outputs a reference signal which serves as a reference signal relative to the detection signal; and a measurement step of determining two candidate phase-change values from a measurement signal which is obtained from the detection signal and the reference signal in accordance with a heterodyne system, and outputting one of the two candidate phase-change values as a phase change value based on temporal changes in the measurement signal, wherein the measurement step comprises mixing the measurement signal and then filtering the mixed measurement signal to filter out higher-frequency components of the mixed measurement signal; an outputting step of outputting a first phase-change value from among the two candidate phase-change values when a strength of the measurement signal is increasing and outputting a second phase-change value from among the two candidate phase-change values when the strength of the measurement signal is decreasing; and a calculation step of calculating an amount of the target in the analyte based on the outputting of the first phase-change value or the second phase-change value.
 22. The analyte sensing method according to claim 21, wherein the temporal changes in measurement signal are obtained by conducting measurement two or more times at time-spaced intervals.
 23. The analyte sensing method according to claim 21, wherein the temporal changes in measurement signal are obtained by conducting continuous measurement.
 24. The analyte sensing method according to claim 21, further comprising a calculation step of calculating a detection amount of the analyte on a basis of the phase change value.
 25. The analyte sensing method according to claim 21, wherein the detection signal output portion comprises an analyte detecting portion which changes in response to adsorption of the target or to reaction with the target.
 26. The analyte sensing method according to claim 25, wherein in the measurement step, a larger one of the two candidate phase-change values is outputted as the phase change value when the temporal changes in measurement signal decrease with time, and a smaller one of the two candidate phase-change values is outputted as the phase change value when the temporal changes in measurement signal increase with time, in a case where a mass of the analyte detecting portion increases and the detection signal is obtained by subtracting the detection signal from the reference signal, or where the mass of the analyte detecting portion decreases and the detection signal is obtained by subtracting the reference signal from the detection signal, and a smaller one of the two candidate phase-change values is outputted as the phase change value when the temporal changes in measurement signal decrease with time, and a larger one of the two candidate phase-change values is outputted as the phase change value when the temporal changes in measurement signal increase with time, in a case where the mass of the analyte detecting portion increases and the detection signal is obtained by subtracting the reference signal from the detection signal, or where the mass of the analyte detecting portion decreases and the detection signal is obtained by subtracting the detection signal from the reference signal.
 27. The analyte sensing method according to claim 21, wherein the two candidate phase-change values are composed of a positive candidate phase-change value and a negative candidate phase-change value.
 28. The analyte sensing method according to claim 21, wherein the measurement step comprises a step of amplifying each of the detection signal and the reference signal, and a step of obtaining the measurement signal on a basis of the detection signal amplified and reference signal amplified, by the heterodyne system.
 29. The analyte sensing method according to claim 21, wherein the reference signal output portion comprises a reference measuring portion which undergoes neither adsorption of the target nor reaction with the target. 